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Fusion of ZMYND8 and RELA genes in acute erythroid leukemia.

Panagopoulos I, Micci F, Thorsen J, Haugom L, Buechner J, Kerndrup G, Tierens A, Zeller B, Heim S - PLoS ONE (2013)

Bottom Line: Cytogenetic analysis of bone marrow (BM) cells showed a t(11;20)(p11;q11) translocation.RT-PCR and direct sequencing verified the presence of an in frame ZMYND8-RELA chimeric transcript.Fluorescence in situ hybridization showed that the ZMYND8-RELA was located on the p12 band of der(11); therefore a cytogenetically invisible pericentric inversion in chromosome 11 must have taken place besides the translocation.

View Article: PubMed Central - PubMed

Affiliation: Section for Cancer Cytogenetics, Institute for Medical Informatics, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway. ioannis.panagopoulos@rr-research.no

ABSTRACT
Acute erythroid leukemia was diagnosed in a 4-month-old boy. Cytogenetic analysis of bone marrow (BM) cells showed a t(11;20)(p11;q11) translocation. RNA extracted from the BM was sequenced and analyzed for fusion transcripts using the software FusionMap. A ZMYND8-RELA fusion was ranked first. RT-PCR and direct sequencing verified the presence of an in frame ZMYND8-RELA chimeric transcript. Fluorescence in situ hybridization showed that the ZMYND8-RELA was located on the p12 band of der(11); therefore a cytogenetically invisible pericentric inversion in chromosome 11 must have taken place besides the translocation. The putative ZMYND8-RELA fusion protein contains the Zinc-PHD finger domain, a bromodomain, a PWWP domain, a MYND type of zinc finger of ZMYND8, and the entire RELA protein, indicating that it might act leukemogenically by influencing several cellular processes including the NF-kappa-B pathway.

Show MeSH

Related in: MedlinePlus

Cytogenetic, FISH, and PCR analyses of the erythroid leukemia.A) Partial G-banded karyotype showing what appeared to be chromosome aberrations der(11)t(11;20)(p11;q13) and der(20)t(11;20)(p11;q13) together with their corresponding normal homologues; breakpoint positions are indicated by arrows. B) FISH using BAC RP11-702E03 (red signal) from 20q13 containing the ZMYND8 gene and BAC CTD-2382C11 (green signal) for RELA. A part of the probe RP11-702E03 for ZMYND8 as well as the entire probe CTD-2382C11 for RELA had moved to band p12 of the derivative chromosome 11. The data suggest that the functional fusion gene is generated on the der(11). C) G-banding of the metaphase spread shown in (B). D) Mapping position of the RP11-702E03 on chromosome band 20q13 and CTD-2382C11 on chromosome band 11q13. E) Amplification of ZMYND8-RELA fusion cDNA fragments using ZMYND8-2683F and RELA-516R primer sets (lane 1) and ZMYND8-3079F and RELA-516R primers (lane 2). M, 1 Kb DNA ladder. F) Partial sequence chromatogram showing the junctions of the ZMYND8-RELA chimeric transcript and part of the in-frame coding protein.
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pone-0063663-g001: Cytogenetic, FISH, and PCR analyses of the erythroid leukemia.A) Partial G-banded karyotype showing what appeared to be chromosome aberrations der(11)t(11;20)(p11;q13) and der(20)t(11;20)(p11;q13) together with their corresponding normal homologues; breakpoint positions are indicated by arrows. B) FISH using BAC RP11-702E03 (red signal) from 20q13 containing the ZMYND8 gene and BAC CTD-2382C11 (green signal) for RELA. A part of the probe RP11-702E03 for ZMYND8 as well as the entire probe CTD-2382C11 for RELA had moved to band p12 of the derivative chromosome 11. The data suggest that the functional fusion gene is generated on the der(11). C) G-banding of the metaphase spread shown in (B). D) Mapping position of the RP11-702E03 on chromosome band 20q13 and CTD-2382C11 on chromosome band 11q13. E) Amplification of ZMYND8-RELA fusion cDNA fragments using ZMYND8-2683F and RELA-516R primer sets (lane 1) and ZMYND8-3079F and RELA-516R primers (lane 2). M, 1 Kb DNA ladder. F) Partial sequence chromatogram showing the junctions of the ZMYND8-RELA chimeric transcript and part of the in-frame coding protein.

Mentions: A 4-month-old boy was referred to hospital with fatigue, irritability, regurgitation, and pallor for the last 3 weeks. On admission, the patient had high fever and hepatosplenomegaly, but otherwise normal physical findings. No congenital abnormalities or dysmorphic features were seen. Initial blood tests showed hemoglobin 6.7 g/dl, white blood cell count 24.9×109/L, platelet count 116×109/L, hemoglobin F 12%, and lactate dehydrogenase 8400 U/L. Peripheral blood smears demonstrated polychromasia and circulating dysplastic erythroblasts of all maturation stages, some metamyelocytes and myelocytes, but no myeloblasts. Examination of a bone marrow (BM) aspirate revealed erythroid hyperplasia and dysplasia with binucleated and multinucleated forms. The aspirate smear count displayed 4% myeloblasts, 38% proerythroblasts, 25% polychromatic erythroblasts, and 22% orthochromatic erythroblasts. Histological examination of the BM revealed high cellularity, dysplastic megakaryocytes, and slight fibrosis around sinusoids. The number of CD34+ cells in the biopsy was around 8%. Flow cytometric immunophenotyping of consecutive bone marrow aspirates showed a marked increase of CD45 negative erytroblasts, ranging from 30-60% of total cells. They expressed CD71, glycophorin, and CD36, but were negative for other cell lineage markers such as CD13, CD33, CD11b, T-and B-cell markers as well as HLA-DR antigens. The CD34+ cells comprised 1.5% of total cells, and were mainly CD34+ B-cell precursor cells. The CD34+ myeloid precursor cells displayed a normal expression of CD34, CD117, HLA-DR antigens, CD13, and CD33. Granulopoiesis revealed a normal maturation pattern as revealed by the expression of CD13 and CD11b. Cytogenetic analysis of BM cells showed the abnormal karyotype 46,XY,t(11;20)(p11;q11) in 8 out of 10 metaphases (Figure 1A and 1C). A myeloid malignancy, either myelodysplastic syndrome or an early phase of AML, possibly with an infection superimposed, was suspected. Prior to the cytogenetics, congenital dyserythropoietic anemia (CDA) was considered and ruled out. Slides of the patient´s bone marrow were also evaluated at the CDA Diagnostic Service and Research Laboratory, MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, Oxord, UK. The morphological appearance was not diagnostic for CDA. During an observational period the child's general condition deteriorated, and hemophagocytic lymphohistiocytosis (HLH) was assumed. Hemophagocytosis was present in the bone marrow. In addition, HLH was suggested due to a combination of clinical criteria (splenomegaly, fever) and the laboratory constellation with cytopenia, hypertriglyceridemia, hyperferritinemia, and neurological symptoms. No infectious agents were detected. Cytology of the cerebrospinal fluid and brain-MRI were normal. Treatment according to an HLH-protocol [9] was initiated. However, repeated BM investigations now demonstrated 100% cellularity and > 70% CD71+ erythroblasts, indicative of a transition to acute erythroid leukemia. The patient did not completely fullfill the criteria “more than 80% erythroblasts” (M6b), nor “more than 20% blasts in the nonerythroid compartment” (M6a) as required by the WHO definition of pure erythroid leukemia [1], which might simply be due to the fact that treatment was started before he reached the appropriate counts. Concurrently, cytogenetic analysis showed clonal evolution to a more complex karyotype: 48,XY,+7,t(11;20)(p11;q11),+17[4]/48,XY,+del(7)(p11)t(11;20)(p11;q11),+17[9]/46,XY[2]. Therefore, HLH-treatment was stopped and AML treatment according to the current Nordic AML protocol begun instead [10]. Morphological bone marrow remission was achieved after the first of two induction courses. After the first consolidation course, the patient was transplanted in cytogenetic remission from a matched unrelated donor after conditioning with busulfan, cyclophosphamide, and melphalan [11]. He is now in complete remission 8 months after the transplantation.


Fusion of ZMYND8 and RELA genes in acute erythroid leukemia.

Panagopoulos I, Micci F, Thorsen J, Haugom L, Buechner J, Kerndrup G, Tierens A, Zeller B, Heim S - PLoS ONE (2013)

Cytogenetic, FISH, and PCR analyses of the erythroid leukemia.A) Partial G-banded karyotype showing what appeared to be chromosome aberrations der(11)t(11;20)(p11;q13) and der(20)t(11;20)(p11;q13) together with their corresponding normal homologues; breakpoint positions are indicated by arrows. B) FISH using BAC RP11-702E03 (red signal) from 20q13 containing the ZMYND8 gene and BAC CTD-2382C11 (green signal) for RELA. A part of the probe RP11-702E03 for ZMYND8 as well as the entire probe CTD-2382C11 for RELA had moved to band p12 of the derivative chromosome 11. The data suggest that the functional fusion gene is generated on the der(11). C) G-banding of the metaphase spread shown in (B). D) Mapping position of the RP11-702E03 on chromosome band 20q13 and CTD-2382C11 on chromosome band 11q13. E) Amplification of ZMYND8-RELA fusion cDNA fragments using ZMYND8-2683F and RELA-516R primer sets (lane 1) and ZMYND8-3079F and RELA-516R primers (lane 2). M, 1 Kb DNA ladder. F) Partial sequence chromatogram showing the junctions of the ZMYND8-RELA chimeric transcript and part of the in-frame coding protein.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3646816&req=5

pone-0063663-g001: Cytogenetic, FISH, and PCR analyses of the erythroid leukemia.A) Partial G-banded karyotype showing what appeared to be chromosome aberrations der(11)t(11;20)(p11;q13) and der(20)t(11;20)(p11;q13) together with their corresponding normal homologues; breakpoint positions are indicated by arrows. B) FISH using BAC RP11-702E03 (red signal) from 20q13 containing the ZMYND8 gene and BAC CTD-2382C11 (green signal) for RELA. A part of the probe RP11-702E03 for ZMYND8 as well as the entire probe CTD-2382C11 for RELA had moved to band p12 of the derivative chromosome 11. The data suggest that the functional fusion gene is generated on the der(11). C) G-banding of the metaphase spread shown in (B). D) Mapping position of the RP11-702E03 on chromosome band 20q13 and CTD-2382C11 on chromosome band 11q13. E) Amplification of ZMYND8-RELA fusion cDNA fragments using ZMYND8-2683F and RELA-516R primer sets (lane 1) and ZMYND8-3079F and RELA-516R primers (lane 2). M, 1 Kb DNA ladder. F) Partial sequence chromatogram showing the junctions of the ZMYND8-RELA chimeric transcript and part of the in-frame coding protein.
Mentions: A 4-month-old boy was referred to hospital with fatigue, irritability, regurgitation, and pallor for the last 3 weeks. On admission, the patient had high fever and hepatosplenomegaly, but otherwise normal physical findings. No congenital abnormalities or dysmorphic features were seen. Initial blood tests showed hemoglobin 6.7 g/dl, white blood cell count 24.9×109/L, platelet count 116×109/L, hemoglobin F 12%, and lactate dehydrogenase 8400 U/L. Peripheral blood smears demonstrated polychromasia and circulating dysplastic erythroblasts of all maturation stages, some metamyelocytes and myelocytes, but no myeloblasts. Examination of a bone marrow (BM) aspirate revealed erythroid hyperplasia and dysplasia with binucleated and multinucleated forms. The aspirate smear count displayed 4% myeloblasts, 38% proerythroblasts, 25% polychromatic erythroblasts, and 22% orthochromatic erythroblasts. Histological examination of the BM revealed high cellularity, dysplastic megakaryocytes, and slight fibrosis around sinusoids. The number of CD34+ cells in the biopsy was around 8%. Flow cytometric immunophenotyping of consecutive bone marrow aspirates showed a marked increase of CD45 negative erytroblasts, ranging from 30-60% of total cells. They expressed CD71, glycophorin, and CD36, but were negative for other cell lineage markers such as CD13, CD33, CD11b, T-and B-cell markers as well as HLA-DR antigens. The CD34+ cells comprised 1.5% of total cells, and were mainly CD34+ B-cell precursor cells. The CD34+ myeloid precursor cells displayed a normal expression of CD34, CD117, HLA-DR antigens, CD13, and CD33. Granulopoiesis revealed a normal maturation pattern as revealed by the expression of CD13 and CD11b. Cytogenetic analysis of BM cells showed the abnormal karyotype 46,XY,t(11;20)(p11;q11) in 8 out of 10 metaphases (Figure 1A and 1C). A myeloid malignancy, either myelodysplastic syndrome or an early phase of AML, possibly with an infection superimposed, was suspected. Prior to the cytogenetics, congenital dyserythropoietic anemia (CDA) was considered and ruled out. Slides of the patient´s bone marrow were also evaluated at the CDA Diagnostic Service and Research Laboratory, MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, Oxord, UK. The morphological appearance was not diagnostic for CDA. During an observational period the child's general condition deteriorated, and hemophagocytic lymphohistiocytosis (HLH) was assumed. Hemophagocytosis was present in the bone marrow. In addition, HLH was suggested due to a combination of clinical criteria (splenomegaly, fever) and the laboratory constellation with cytopenia, hypertriglyceridemia, hyperferritinemia, and neurological symptoms. No infectious agents were detected. Cytology of the cerebrospinal fluid and brain-MRI were normal. Treatment according to an HLH-protocol [9] was initiated. However, repeated BM investigations now demonstrated 100% cellularity and > 70% CD71+ erythroblasts, indicative of a transition to acute erythroid leukemia. The patient did not completely fullfill the criteria “more than 80% erythroblasts” (M6b), nor “more than 20% blasts in the nonerythroid compartment” (M6a) as required by the WHO definition of pure erythroid leukemia [1], which might simply be due to the fact that treatment was started before he reached the appropriate counts. Concurrently, cytogenetic analysis showed clonal evolution to a more complex karyotype: 48,XY,+7,t(11;20)(p11;q11),+17[4]/48,XY,+del(7)(p11)t(11;20)(p11;q11),+17[9]/46,XY[2]. Therefore, HLH-treatment was stopped and AML treatment according to the current Nordic AML protocol begun instead [10]. Morphological bone marrow remission was achieved after the first of two induction courses. After the first consolidation course, the patient was transplanted in cytogenetic remission from a matched unrelated donor after conditioning with busulfan, cyclophosphamide, and melphalan [11]. He is now in complete remission 8 months after the transplantation.

Bottom Line: Cytogenetic analysis of bone marrow (BM) cells showed a t(11;20)(p11;q11) translocation.RT-PCR and direct sequencing verified the presence of an in frame ZMYND8-RELA chimeric transcript.Fluorescence in situ hybridization showed that the ZMYND8-RELA was located on the p12 band of der(11); therefore a cytogenetically invisible pericentric inversion in chromosome 11 must have taken place besides the translocation.

View Article: PubMed Central - PubMed

Affiliation: Section for Cancer Cytogenetics, Institute for Medical Informatics, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway. ioannis.panagopoulos@rr-research.no

ABSTRACT
Acute erythroid leukemia was diagnosed in a 4-month-old boy. Cytogenetic analysis of bone marrow (BM) cells showed a t(11;20)(p11;q11) translocation. RNA extracted from the BM was sequenced and analyzed for fusion transcripts using the software FusionMap. A ZMYND8-RELA fusion was ranked first. RT-PCR and direct sequencing verified the presence of an in frame ZMYND8-RELA chimeric transcript. Fluorescence in situ hybridization showed that the ZMYND8-RELA was located on the p12 band of der(11); therefore a cytogenetically invisible pericentric inversion in chromosome 11 must have taken place besides the translocation. The putative ZMYND8-RELA fusion protein contains the Zinc-PHD finger domain, a bromodomain, a PWWP domain, a MYND type of zinc finger of ZMYND8, and the entire RELA protein, indicating that it might act leukemogenically by influencing several cellular processes including the NF-kappa-B pathway.

Show MeSH
Related in: MedlinePlus